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Showing papers by "Matthias Meyer published in 2018"


Journal ArticleDOI
22 Aug 2018-Nature
TL;DR: The genome of ‘Denisova 11’, a bone fragment from Denisova Cave (Russia) is presented and it is shown that it comes from an individual who had a Neanderthal mother and a Denisovan father.
Abstract: Neanderthals and Denisovans are extinct groups of hominins that separated from each other more than 390,000 years ago1,2. Here we present the genome of ‘Denisova 11’, a bone fragment from Denisova Cave (Russia)3 and show that it comes from an individual who had a Neanderthal mother and a Denisovan father. The father, whose genome bears traces of Neanderthal ancestry, came from a population related to a later Denisovan found in the cave4–6. The mother came from a population more closely related to Neanderthals who lived later in Europe2,7 than to an earlier Neanderthal found in Denisova Cave8, suggesting that migrations of Neanderthals between eastern and western Eurasia occurred sometime after 120,000 years ago. The finding of a first-generation Neanderthal–Denisovan offspring among the small number of archaic specimens sequenced to date suggests that mixing between Late Pleistocene hominin groups was common when they met.

274 citations


Journal ArticleDOI
21 Mar 2018-Nature
TL;DR: It is found that the bulk of Neanderthal gene flow into early modern humans originated from one or more source populations that diverged from the Neanderthals that were studied here at least 70,000 years ago, but after they split from a previously sequenced Neanderthal from Siberia around 150,000 year ago.
Abstract: Although it has previously been shown that Neanderthals contributed DNA to modern humans, not much is known about the genetic diversity of Neanderthals or the relationship between late Neanderthal populations at the time at which their last interactions with early modern humans occurred and before they eventually disappeared. Our ability to retrieve DNA from a larger number of Neanderthal individuals has been limited by poor preservation of endogenous DNA and contamination of Neanderthal skeletal remains by large amounts of microbial and present-day human DNA. Here we use hypochlorite treatment of as little as 9 mg of bone or tooth powder to generate between 1- and 2.7-fold genomic coverage of five Neanderthals who lived around 39,000 to 47,000 years ago (that is, late Neanderthals), thereby doubling the number of Neanderthals for which genome sequences are available. Genetic similarity among late Neanderthals is well predicted by their geographical location, and comparison to the genome of an older Neanderthal from the Caucasus indicates that a population turnover is likely to have occurred, either in the Caucasus or throughout Europe, towards the end of Neanderthal history. We find that the bulk of Neanderthal gene flow into early modern humans originated from one or more source populations that diverged from the Neanderthals that were studied here at least 70,000 years ago, but after they split from a previously sequenced Neanderthal from Siberia around 150,000 years ago. Although four of the Neanderthals studied here post-date the putative arrival of early modern humans into Europe, we do not detect any recent gene flow from early modern humans in their ancestry.

188 citations


Journal ArticleDOI
TL;DR: This protocol update replaces a now-outdated version that was published 11 years ago, before high-throughput sequencing technologies became widely available, and has been thoroughly optimized to provide the highest DNA yields from highly degraded samples, as well as fast and easy handling.
Abstract: DNA preserved in ancient bones, teeth and sediments is typically highly fragmented and present only in minute amounts. Here, we provide a highly versatile silica-based DNA extraction protocol that enables the retrieval of short (≥35 bp) or even ultrashort (≥25 bp) DNA fragments from such material with minimal carryover of substances that inhibit library preparation for high-throughput sequencing. DNA extraction can be performed with either silica spin columns, which offer the most convenient choice for manual DNA extraction, or silica-coated magnetic particles. The latter allow a substantial cost reduction as well as automation on liquid-handling systems. This protocol update replaces a now-outdated version that was published 11 years ago, before high-throughput sequencing technologies became widely available. It has been thoroughly optimized to provide the highest DNA yields from highly degraded samples, as well as fast and easy handling, requiring not more than ~15 min of hands-on time per sample. This protocol update describes silica-based approaches for purification of DNA from ancient bone, tooth and sediment samples. The optimized buffers yield short DNA fragments compatible with high-throughput sequencing library preparation.

144 citations


Journal ArticleDOI
04 May 2018-Science
TL;DR: Genomic data from seven 15,000-year-old modern humans attributed to the Iberomaurusian culture from Morocco are presented, finding a genetic affinity with early Holocene Near Easterners, best represented by Levantine Natufians, suggesting a pre-agricultural connection between Africa and the Near East.
Abstract: North Africa is a key region for understanding human history, but the genetic history of its people is largely unknown. We present genomic data from seven 15,000-year-old modern humans, attributed to the Iberomaurusian culture, from Morocco. We find a genetic affinity with early Holocene Near Easterners, best represented by Levantine Natufians, suggesting a pre-agricultural connection between Africa and the Near East. We do not find evidence for gene flow from Paleolithic Europeans to Late Pleistocene North Africans. The Taforalt individuals derive one-third of their ancestry from sub-Saharan Africans, best approximated by a mixture of genetic components preserved in present-day West and East Africans. Thus, we provide direct evidence for genetic interactions between modern humans across Africa and Eurasia in the Pleistocene.

123 citations


Posted ContentDOI
Vagheesh M. Narasimhan1, Nick Patterson2, Nick Patterson3, Priya Moorjani4, Iosif Lazaridis1, Mark Lipson1, Swapan Mallick5, Swapan Mallick3, Nadin Rohland3, Rebecca Bernardos1, Alexander M. Kim1, Nathan Nakatsuka6, Iñigo Olalde1, Alfredo Coppa7, James Mallory8, Vyacheslav Moiseyev9, Janet Monge10, Luca M. Olivieri, Nicole Adamski5, Nasreen Broomandkhoshbacht5, Francesca Candilio10, Olivia Cheronet11, Olivia Cheronet12, Brendan J. Culleton13, Matthew Ferry5, Daniel Fernandes, Beatriz Gamarra11, Daniel Gaudio11, Mateja Hajdinjak14, Eadaoin Harney5, Thomas K. Harper13, Denise Keating11, Ann Marie Lawson5, Megan Michel5, Mario Novak11, Jonas Oppenheimer5, Niraj Rai15, Niraj Rai16, Kendra Sirak17, Kendra Sirak1, Kendra Sirak11, Viviane Slon14, Kristin Stewardson5, Zhao Zhang1, Gaziz Akhatov, Anatoly N. Bagashev, Bauryzhan Baitanayev, Gian Luca Bonora, T. A. Chikisheva9, Anatoly P. Derevianko9, Dmitry Enshin, Katerina Douka14, Katerina Douka18, Nadezhda Dubova9, Andrey Epimakhov19, Suzanne Freilich12, Dorian Q. Fuller20, Dorian Q. Fuller21, Alexander Goryachev, Andrey Gromov9, Bryan Hanks22, Margaret A. Judd22, Erlan Kazizov, Aleksander Khokhlov23, Egor Kitov9, Egor Kitov24, Elena Kupriyanova25, Pavel Kuznetsov23, Donata Luiselli26, Farhod Maksudov27, Christopher Meiklejohn28, Deborah C. Merrett29, Roberto Micheli, Oleg Mochalov23, Muhammad Zahir30, Muhammad Zahir14, Samariddin Mustafokulov27, Ayushi Nayak14, Marina Petrovna Rykun31, Davide Pettener26, Richard Potts32, Dmitry Razhev, Stefania Sarno26, Kulyan Sikhymbaeva, Sergey Mikhailovich Slepchenko, Nadezhda Stepanova9, Svetlana V. Svyatko9, Svetlana V. Svyatko8, Sergey Vasilyev9, Massimo Vidale33, Dmitriy Voyakin, Antonina Yermolayeva, A. V. Zubova9, Vasant Shinde34, Carles Lalueza-Fox35, Matthias Meyer14, David W. Anthony36, Nicole Boivin14, Kumarasamy Thangaraj15, Douglas J. Kennett37, Douglas J. Kennett13, Michael D. Frachetti38, Ron Pinhasi11, Ron Pinhasi12, David Reich 
31 Mar 2018-bioRxiv
TL;DR: The results show how ancestry from the Steppe genetically linked Europe and South Asia in the Bronze Age, and identifies the populations that almost certainly were responsible for spreading Indo-European languages across much of Eurasia.
Abstract: The genetic formation of Central and South Asian populations has been unclear because of an absence of ancient DNA. To address this gap, we generated genome-wide data from 362 ancient individuals, including the first from eastern Iran, Turan (Uzbekistan, Turkmenistan, and Tajikistan), Bronze Age Kazakhstan, and South Asia. Our data reveal a complex set of genetic sources that ultimately combined to form the ancestry of South Asians today. We document a southward spread of genetic ancestry from the Eurasian Steppe, correlating with the archaeologically known expansion of pastoralist sites from the Steppe to Turan in the Middle Bronze Age (2300-1500 BCE). These Steppe communities mixed genetically with peoples of the Bactria Margiana Archaeological Complex (BMAC) whom they encountered in Turan (primarily descendants of earlier agriculturalists of Iran), but there is no evidence that the main BMAC population contributed genetically to later South Asians. Instead, Steppe communities integrated farther south throughout the 2nd millennium BCE, and we show that they mixed with a more southern population that we document at multiple sites as outlier individuals exhibiting a distinctive mixture of ancestry related to Iranian agriculturalists and South Asian hunter-gathers. We call this group Indus Periphery because they were found at sites in cultural contact with the Indus Valley Civilization (IVC) and along its northern fringe, and also because they were genetically similar to post-IVC groups in the Swat Valley of Pakistan. By co-analyzing ancient DNA and genomic data from diverse present-day South Asians, we show that Indus Periphery-related people are the single most important source of ancestry in South Asia—consistent with the idea that the Indus Periphery individuals are providing us with the first direct look at the ancestry of peoples of the IVC—and we develop a model for the formation of present-day South Asians in terms of the temporally and geographically proximate sources of Indus Periphery-related, Steppe, and local South Asian hunter-gatherer-related ancestry. Our results show how ancestry from the Steppe genetically linked Europe and South Asia in the Bronze Age, and identifies the populations that almost certainly were responsible for spreading Indo-European languages across much of Eurasia. One Sentence Summary Genome wide ancient DNA from 357 individuals from Central and South Asia sheds new light on the spread of Indo-European languages and parallels between the genetic history of two sub-continents, Europe and South Asia.

61 citations


Journal ArticleDOI
TL;DR: Combined DNA and collagen extraction not only streamlines the sampling process but also drastically increases the amount of DNA that can be recovered from limited sample material.
Abstract: Current protocols for ancient DNA and radiocarbon analysis of ancient bones and teeth call for multiple destructive samplings of a given specimen, thereby increasing the extent of undesirable damage to precious archaeological material. Here we present a method that makes it possible to obtain both ancient DNA sequences and radiocarbon dates from the same sample material. This is achieved by releasing DNA from the bone matrix through incubation with either EDTA or phosphate buffer prior to complete demineralization and collagen extraction utilizing the acid-base-acid-gelatinization and ultrafiltration procedure established in most radiocarbon dating laboratories. Using a set of 12 bones of different ages and preservation conditions we demonstrate that on average 89% of the DNA can be released from sample powder with minimal, or 38% without any, detectable collagen loss. We also detect no skews in radiocarbon dates compared to untreated samples. Given the different material demands for radiocarbon dating (500 mg of bone/dentine) and DNA analysis (10–100 mg), combined DNA and collagen extraction not only streamlines the sampling process but also drastically increases the amount of DNA that can be recovered from limited sample material.

40 citations


Journal ArticleDOI
TL;DR: A method to quantify spurious alignments based on the presence or absence of rare variants is developed, which maximizes the sequence data amenable to genetic analysis from highly degraded ancient material and avoids pitfalls that are associated with the analysis of ultra-short DNA sequences.
Abstract: The study of ancient DNA is hampered by degradation, resulting in short DNA fragments. Advances in laboratory methods have made it possible to retrieve short DNA fragments, thereby improving access to DNA preserved in highly degraded, ancient material. However, such material contains large amounts of microbial contamination in addition to DNA fragments from the ancient organism. The resulting mixture of sequences constitutes a challenge for computational analysis, since microbial sequences are hard to distinguish from the ancient sequences of interest, especially when they are short. Here, we develop a method to quantify spurious alignments based on the presence or absence of rare variants. We find that spurious alignments are enriched for mismatches and insertion/deletion differences and lack substitution patterns typical of ancient DNA. The impact of spurious alignments can be reduced by filtering on these features and by imposing a sample-specific minimum length cutoff. We apply this approach to sequences from four ~ 430,000-year-old Sima de los Huesos hominin remains, which contain particularly short DNA fragments, and increase the amount of usable sequence data by 17–150%. This allows us to place a third specimen from the site on the Neandertal lineage. Our method maximizes the sequence data amenable to genetic analysis from highly degraded ancient material and avoids pitfalls that are associated with the analysis of ultra-short DNA sequences.

38 citations


Posted ContentDOI
Vagheesh M. Narasimhan, Nick Patterson1, Priya Moorjani2, Iosif Lazaridis2, Lipson Mark3, Swapan Mallick4, Nadin Rohland2, Rebecca Bernardos1, Alexander M. Kim, Nathan Nakatsuka, Iñigo Olalde, Alfredo Coppa, James Mallory, Vyacheslav Moiseyev, Janet Monge, Luca M. Olivieri, Nicole Adamski, Nasreen Broomandkhoshbacht, Francesca Candilio, Olivia Cheronet, Brendan J. Culleton, Matthew Ferry, Daniel Fernandes, Beatriz Gamarra, Daniel Gaudio, Mateja Hajdinjak, Eadaoin Harney, Thomas K. Harper, Denise Keating, Ann-Marie Lawson, Megan Michel, Mario Novak, Jonas Oppenheimer, Niraj Rai, Kendra Sirak, Viviane Slon, Kristin Stewardson, Zhao Zhang, Gaziz Akhatov, Anatoly N. Bagashev, Baurzhan Baitanayev, Gian Luca Bonora, T. A. Chikisheva, Anatoly P. Derevianko, Enshin Dmitry, Katerina Douka3, Nadezhda Dubova, Andrey Epimakhov, Suzanne Freilich, Dorian Q. Fuller, Alexander Goryachev, Andrey Gromov, Bryan Hanks, Margaret A. Judd, Erlan Kazizov, Aleksander Khokhlov, Egor Kitov, Elena Kupriyanova, Pavel Kuznetsov, Donata Luiselli, Farhad Maksudov, Christopher Meiklejohn, Deborah C. Merrett, Roberto Micheli, Oleg Mochalov, Muhammad Zahir3, Samridin Mustafakulov, Ayushi Nayak3, Rykun M Petrovna, Davide Pettner, Richard Potts, Dmitry Razhev, Stefania Sarno, Kulyan Sikhymbaevae, Sergey M. Slepchenko, Nadezhda Stepanova, Svetlana V. Svyatko, Sergey Vasilyev, Massimo Vidale, Dima Voyakin, Antonina Yermolayeva, A. V. Zubova, Vasant Shinde, Carles Lalueza-Fox, Matthias Meyer, David W. Anthony, Nicole Boivin3, Kumarasmy Thangaraj4, Douglas J. Kennett, Michael D. Frachetti, Ron Pinhasi, David Reich1 
31 Mar 2018-Science
TL;DR: The Steppe ancestry in South Asia has the same profile as that in Bronze Age Eastern Europe, tracking a movement of people that affected both regions and that likely spread the distinctive features shared between Indo-Iranian and Balto-Slavic languages.
Abstract: By sequencing 523 ancient humans, we show that the primary source of ancestry in modern South Asians is a prehistoric genetic gradient between people related to early hunter-gatherers of Iran and Southeast Asia. After the Indus Valley Civilization’s decline, its people mixed with individuals in the southeast to form one of the two main ancestral populations of South Asia, whose direct descendants live in southern India. Simultaneously, they mixed with descendants of Steppe pastoralists who, starting around 4000 years ago, spread via Central Asia to form the other main ancestral population.The Steppe ancestry in South Asia has the same profile as that in Bronze Age Eastern Europe, tracking a movement of people that affected both regions and that likely spread the distinctive features shared between Indo-Iranian and Balto-Slavic languages.

32 citations


Journal ArticleDOI
TL;DR: In this article, the authors used the Roche SeqCap EZ developer kit to capture a complex pool of DNA libraries for exome exome capture and compared the effect of using more than one fecal sample, extract and/or library from the same individual.
Abstract: Target capture approach has improved over the past years, proving to be very efficient tool for selectively sequencing genetic regions of interest. These methods have also allowed the use of non-invasive samples such as feces (characterized by their low quantity and quality of endogenous DNA) to be used in conservation genomic, evolution, and population genetic studies. Here we aim to test different protocols and strategies for exome capture using the Roche SeqCap EZ Developer kit (57.5 Mb). First, we captured a complex pool of DNA libraries. Second, we assessed the influence of using more than one fecal sample, extract and/or library from the same individual, to evaluate its effect on the molecular complexity of the experiment. We validated our experiments with 18 chimpanzee fecal samples collected from two field sites as a part of the Pan African Programme: The Cultured Chimpanzee. Those two field sites are in Kibale National Park, Uganda (N=9) and Loango National Park, Gabon (N=9). We demonstrate that at least 16 libraries can be pooled, target enriched through hybridization, and sequenced allowing for the genotyping of 951949 exome markers for population genetic analyses. Further, we observe that molecule richness, and thus, data acquisition, increases when using multiple libraries from the same extract or multiple extracts from the same sample. Finally, repeated captures significantly decreases the proportion of off-target reads from 34.15% after one capture round to 7.83% after two capture rounds, supporting our conclusion that two rounds of target enrichment are advisable when using complex fecal samples. This article is protected by copyright. All rights reserved.

31 citations


Journal ArticleDOI
TL;DR: The phylogenetic placement of the ancient European leopard as sister group to Asian leopards suggests that these populations originate from the same out-of-Africa dispersal which founded the Asian lineages.
Abstract: Resolving the historical biogeography of the leopard (Panthera pardus) is a complex issue, because patterns inferred from fossils and from molecular data lack congruence. Fossil evidence supports an African origin, and suggests that leopards were already present in Eurasia during the Early Pleistocene. Analysis of DNA sequences however, suggests a more recent, Middle Pleistocene shared ancestry of Asian and African leopards. These contrasting patterns led researchers to propose a two-stage hypothesis of leopard dispersal out of Africa: an initial Early Pleistocene colonisation of Asia and a subsequent replacement by a second colonisation wave during the Middle Pleistocene. The status of Late Pleistocene European leopards within this scenario is unclear: were these populations remnants of the first dispersal, or do the last surviving European leopards share more recent ancestry with their African counterparts? In this study, we generate and analyse mitogenome sequences from historical samples that span the entire modern leopard distribution, as well as from Late Pleistocene remains. We find a deep bifurcation between African and Eurasian mitochondrial lineages (~ 710 Ka), with the European ancient samples as sister to all Asian lineages (~ 483 Ka). The modern and historical mainland Asian lineages share a relatively recent common ancestor (~ 122 Ka), and we find one Javan sample nested within these. The phylogenetic placement of the ancient European leopard as sister group to Asian leopards suggests that these populations originate from the same out-of-Africa dispersal which founded the Asian lineages. The coalescence time found for the mitochondrial lineages aligns well with the earliest undisputed fossils in Eurasia, and thus encourages a re-evaluation of the identification of the much older putative leopard fossils from the region. The relatively recent ancestry of all mainland Asian leopard lineages suggests that these populations underwent a severe population bottleneck during the Pleistocene. Finally, although only based on a single sample, the unexpected phylogenetic placement of the Javan leopard could be interpreted as evidence for exchange of mitochondrial lineages between Java and mainland Asia, calling for further investigation into the evolutionary history of this subspecies.

15 citations


Journal ArticleDOI
TL;DR: Monte Carlo ray-tracing modeling is used to follow the stochastic trajectories of rays entering a cylindrical port from inside an integrating sphere to study and quantify properties of realistic ports of non-negligible length.
Abstract: We use Monte Carlo ray-tracing modeling to follow the stochastic trajectories of rays entering a cylindrical port from inside an integrating sphere. This allows us to study and quantify properties of realistic ports of non-negligible length, as opposed to the common thin-port assumption used in most theoretical treatments, where the port is simply considered as a hole in the spherical wall. We show that most practical ports encountered in integrating sphere applications cannot be modeled as thin ports. Indeed, a substantial proportion of rays entering the port can be reflected back into the sphere, with port reflectances as high as 80% demonstrated on realistic examples. This can have significant consequences on estimates of the sphere multiplier and therefore pathlength inside the sphere, a critical parameter in many applications. Moreover, a nonzero port reflectance is inevitably associated with reduced transmittance through the port, with implications in terms of overall throughput. We also discuss angular redistribution effects in a realistic port and the consequences in terms of detected throughput within a fixed numerical aperture. Those results highlight the importance of real port effects for any quantitative predictions of optical systems using integrating spheres. We believe that those effects can be exploited to engineer ports for specific applications and improve the overall sphere performance in terms of pathlength or throughput. This work carries important implications in our theoretical understanding of integrating spheres and on the practical design of optical systems using them.

Posted ContentDOI
22 Aug 2018-bioRxiv
TL;DR: This work employs a library preparation method that separates molecules that carry uracils from those that do not in a set of samples that includes Neandertal remains, herbaria specimens and archaeological plant remains and shows that this method facilitates the discovery of authentic ancient microbial taxa.
Abstract: The identification of bona fide microbial taxa in microbiomes derived from historic samples is complicated by the unavoidable mixture between DNA from ante- and post-mortem microbial colonizers. One possibility to distinguish between these sources of microbial DNA is querying for the presence of age-associated degradation patterns typical of ancient DNA (aDNA). The presence of uracils, resulting from cytosine deamination, has been detected ubiquitously in aDNA retrieved from diverse sources, and used as an authentication criterion. Here, we employ a library preparation method that separates molecules that carry uracils from those that do not in a set of samples that includes Neandertal remains, herbaria specimens and archaeological plant remains. We show that this method facilitates the discovery of authentic ancient microbial taxa, as it amplifies degradation patterns that would otherwise be difficult to detect in sequences from diverse microbial mixtures.

Posted ContentDOI
10 May 2018-bioRxiv
TL;DR: This work develops a method to quantify spurious alignments based on the presence or absence of rare variants and finds that spuriousalignments are enriched for mismatches and insertion/deletion differences and lack substitution patterns typical of ancient DNA.
Abstract: The study of ancient DNA is hampered by degradation, resulting in short DNA fragments. Advances in laboratory methods have made it possible to retrieve short DNA fragments, thereby improving access to DNA preserved in highly degraded, ancient material. However, such material contains large amounts of microbial contamination in addition to DNA fragments from the ancient organism. The resulting mixture of sequences constitute a challenge for computational analysis, since microbial sequences are hard to distinguish from the ancient sequences of interest, especially when they are short. Here, we develop a method to quantify spurious alignments based on the presence or absence of rare variants. We find that spurious alignments are enriched for mismatches and insertion/deletion differences and lack substitution patterns typical of ancient DNA. The impact of spurious alignments can be reduced by filtering on these features and by imposing a sample-specific minimum length cutoff. We apply this approach to sequences from the ~430,000 year-old Sima de los Huesos hominin remains, which contain particularly short DNA fragments, and increase the amount of usable sequence data by 17-150%. This allows us to place a third specimen from the site on the Neandertal lineage. Our method maximizes the sequence data amenable to genetic analysis from highly degraded ancient material and avoids pitfalls that are associated with the analysis of ultra-short DNA sequences.

Posted ContentDOI
12 Sep 2018-bioRxiv
TL;DR: The phylogenetic placement of the ancient European leopard as sister group to Asian leopards suggests that these populations originate from the same out-of-Africa dispersal which founded the Asian lineages.
Abstract: Background: Resolving the historical biogeography of the leopard (Panthera pardus) is a complex issue, because patterns inferred from fossils and from molecular data lack congruence. Fossil evidence supports an African origin, and suggests that leopards were already present in Eurasia during the Early Pleistocene. Analysis of DNA sequences however, suggests a more recent, Middle Pleistocene shared ancestry of Asian and African leopards. These contrasting patterns led researchers to propose a two-stage hypothesis of leopard dispersal out of Africa: an initial Early Pleistocene colonisation of Asia and a subsequent replacement by a second colonisation wave during the Middle Pleistocene. The status of Late Pleistocene European leopards within this scenario is unclear: were these populations remnants of the first dispersal, or do the last surviving European leopards share more recent ancestry with their African counterparts? Results: In this study, we generate and analyse mitogenome sequences from historical samples that span the entire modern leopard distribution, as well as from Late Pleistocene remains. We find a deep bifurcation between African and Eurasian mitochondrial lineages (~710 Ka), with the European ancient samples as sister to all Asian lineages (~483 Ka). The modern and historical mainland Asian lineages share a relatively recent common ancestor (~122 Ka), and we find one Javan sample nested within these. Conclusions: The phylogenetic placement of the ancient European leopard as sister group to Asian leopards suggests that these populations originate from the same out-of-Africa dispersal which founded the Asian lineages. The coalescence time found for the mitochondrial lineages aligns well with the earliest undisputed fossils in Eurasia, and thus encourages a re-evaluation of the identification of the much older putative leopard fossils from the region. The relatively recent ancestry of all mainland Asian leopard lineages suggests that these populations underwent a severe population bottleneck during the Pleistocene. Finally, although only based on a single sample, the unexpected phylogenetic placement of the Javan leopard could be interpreted as evidence for exchange of mitochondrial lineages between Java and mainland Asia, calling for further investigation into the evolutionary history of this subspecies.